Response of Hydraulic and Photosynthetic Characteristics of Caroxylon passerinum (Bunge) Akhani and Roalson to Prolonged Drought and Short-Term Rehydration

Abstract

Hydraulic traits are essential functional characteristics of plants related to water absorption, transport, and loss, serving as indicators of a plant’s adaptability to prevailing environmental water conditions. However, the hydraulic traits of shrub, particularly desert plants in arid and semi-arid regions, have been underexplored. In this study, we conducted a pot experiment using Caroxylon passerinum (Bunge) Akhani and Roalson as the subject. Three treatment groups were established: adequate water supply, mild drought, and severe drought. After subjecting the shrub to drought and subsequent rehydration, we measured hydraulic conductivity, net photosynthetic rate, and stomatal conductance. We found that leaf water potential decreased and stomatal conductance and net photosynthesis decreased with increasing drought intensity. We found that leaf water potential and stomatal conductance decreased with increasing drought intensity. Although there was no significant change in hydraulic conductivity in the two drought groups, the values were greater in the drought group than in the control, and greater in the mild drought group than in the severe drought group. Meanwhile, the embolism resistance decreased with increasing drought intensity. After rehydration, hydraulic conductivity did not return to control levels in the severe drought group, as did embolism resistance in the two drought groups, and leaf water potential did not recover significantly. The results showed that drought stress increased the hydraulic conductivity of C. passerinum, and this effect was more pronounced under mild drought stress. After the stress was lifted, C. passerinum continued to maintain a lower leaf water potential to promote water uptake. This result provides a reference for us to study water use of desert shrubs under different drought stresses.

1. Introduction

In recent years, global warming has resulted in more frequent drought events [1,2], posing a threat to forest productivity in many regions [3]. The frequency, duration, and severity of droughts are projected to increase significantly in most parts of the globe [4]. Therefore, we urgently need to understand the stress response of shrub to drought, which is a prerequisite for shrub to adapt to climate change.

The water transport system of woody plants efficiently moves water from the soil to various organs, ensuring the regular supply of water to the plant [5]. This system comprises two essential components: xylem hydraulic conductivity, which represents the effectiveness of water transport, and resistance to embolism, which reflects the safety of the transport process [6,7]. Usually, sapwood-specific conductance and leaf-specific conductance are employed to quantify the hydraulic conductivity of plants, as these parameters are closely associated with leaf transpiration and photosynthesis [5,6,8]. The water potential value of hydraulic conductivity at 50% loss of xylem has been used to quantify the plant’s resistance to embolism, with more negative values indicating higher resistance in the plant species [6]. Currently, research endeavors in hydraulic traits of woody plants are concentrated in tropical and subtropical regions [9,10], and there is very little information about hydraulic traits of shrub species in arid and semi-arid regions.

When drought stress is alleviated, the ability of the water transport process in the shrub to quickly return to its pre-stress state becomes an important criterion for assessing the plant’s level of drought tolerance [11]. Numerous studies have shown that the repair time of embolized xylem is shorter under mild drought stress but longer under extreme drought stress [12,13]. This indicates that the repair time of xylem embolism is correlated with the severity of drought stress. Therefore, exploring the changes in hydraulic traits of woody plants after rehydration is important for plant recovery after drought stress is lifted [14].

In this study, Caroxylon passerinum (Bunge) Akhani and Roalson was chosen as the test material. C. passerinum is a significant foundational species in plant communities found in semi-arid and arid regions, and it has shown strong drought resistance due to long-term environmental selection [15]. During our investigation, we examined several physiological traits related to water and photosynthesis under drought stress and subsequent recovery. Our objectives were (1) to analyze changes in the hydraulic and photosynthetic characteristics of C. passerinum under mild and severe drought stress, and (2) to investigate whether water-use efficiency of C. passerinum under drought stress recovered after rehydration.

2. Materials and Methods

2.1. Plant Material

The experiment was conducted at the campus of Gansu Agricultural University. On 25 March 2021, several Caroxylon passerinum (Bunge) Akhani and Roalson Seedlings of uniform size and vigor were selected from natural habitats with an average annual rainfall of 184.8 mm and a canopy spread of approximately 10 cm. Seedlings measuring approximately 10 cm in height and 10 cm² in crown width were chosen from natural habitats characterized by an average annual precipitation of 184.8 mm. These seedlings were then transplanted into pots with an upper diameter of 30 cm, a lower diameter of 18 cm, and a height of 38 cm. Each pot accommodated a single plant, and approximately 10 kg of a culture medium, consisting of a 4:1 blend of native soil and perlite, was added to each pot. Finally, the potted seedlings were positioned under a rain shelter.

2.2. Experimental Design

The treatments began on 10 July 2021, and we randomly assigned C. passerinum seedlings to three water treatments: control, mild drought, and severe drought. We employed a weighed water control method to regulate the soil water content of all plants every two days, considering the variations in water consumption among individual plants. The soil water content required for normal plant growth was measured to be 9.5% before the start of the experiment. Consequently, individuals in the adequately watered group were consistently maintained at 9.5% soil water content throughout the experimental period. Under the mild drought treatment, individuals were rewatered to replenish 60% of the water loss each time. Conversely, individuals subjected to the severe drought treatment received no additional watering. Six plants were treated under each of the three water gradients, for a total of 18 shrubs. The hydraulic and photosynthetic characteristics of the plants were measured after 45 days of continuous water treatment (mean plant height 31.11 cm, base diameter 3.81 mm and crown spread 150.47 cm²). Afterwards, the remaining plants were rehydrated to achieve a soil water content of 9.5%. After 15 days, the same indicators mentioned above were measured in the same manner (

Table 1. Abbreviations and definitions of traits measured during the drought study and rehydration.

Acronym	Definition	Units
ΨPD	Predawn water potential	MPa
ΨMD	Midday water potential	MPa
Pn	Net photosynthetic rate	μmol·m−2·s−1
Tr	Transpiration rate	mmol·m−2·s−1
Gs	Stomatal conductance	mmol·m−2·s−1
WUE	Water use efficiency	μmol/mmol−1
Kh	Natural hydraulic conductivity	kg·m−1·s−1·MPa−1
KS	Sapwood-specific conductivity	kg·m−1·s−1·MPa−1
KL	Leaf-specific conductivity	kg·m−1·s−1·MPa−1
P50	Pressure value in xylem at 50% loss of hydraulic conductivity	MPa

).

2.3. Leaf Water Potential and Photosynthetic Characteristics

Two time points, predawn (4:00–6:00) and midday (12:00–14:00), were chosen to measure leaf water potential representing the maximum and minimum leaf water potential of the plants throughout the day, technical abbreviations are explained. Specifically, the upper 10 cm of leafy tops of single plants of seedlings in the same pot were cut and subsequently measured using a pressure chamber (Model 1505D; PMS Instrument Company, Albany, OR, USA). Each plant was measured 3 times (n = 9). Before measuring the leaf water potential, we chose a sunny day to measure the net photosynthetic rate, transpiration rate and stomatal conductance of living plants in the natural state using a plant photosynthesis meter (3051D, Zhejiang Top Instrument Co., Ltd., Hangzhou, China), and calculate the instantaneous water use efficiency (WUE) in terms of Pn/Tr.

2.4. Water Conductivity and Susceptibility to Embolism

Determination of hydraulic conductivity requires that the length of the branch be greater than the maximum conduit length. In our study, we measured 7–8 branches using the air injection method [16] and calculated the average as the maximum conduit length, which was found to be 7.8 cm. Therefore, we used 20 cm long C. passerinum branches for the assay. First, we cut 30 cm branches, sealed them by wrapping them in a wet towel, and promptly transferred them to a cooler. The branches were then brought back to the laboratory and processed at a controlled temperature of 20 °C–25 °C. The branch ends were recut by 2–3 cm in distilled water. Subsequently, the measurement of stem xylem hydraulic conductivity was conducted using the low-pressure liquid flow technique [17]. Measurements were taken from 1 branch per plant. There were 3 replicates per treatment.

The branches were connected to the hydraulic conductivity device, and water flow was induced through the branches using a water purification pressure of 5 KPa. A point to note is that the injection stream was consistently directed from the bottom to the top of the stem. The amount of water passing through the branches over 20 min was measured to determine the natural hydraulic conductivity (K_h, kg·m·s⁻¹·Mpa⁻¹). Subsequently, the xylem of the branches was rinsed with a pressure of 200 kPa for 20 min, and the hydraulic conductivity of the xylem was measured under a pressure of 5 kPa to determine the maximum hydraulic conductivity (K_max, kg·m·s⁻¹·Mpa⁻¹).

Following the aforementioned measurements, the branches were positioned within pressure chambers (PMS, Corvallis, OR, USA) extending from both ends. A total of 12 sequential pressurizations were carried out via the chambers to create stem xylem embolism fragility curves. These curves were established by recording the hydraulic conductivity loss values corresponding to specific pressures and durations, each lasting 20 min. The embolic fragility curves were fitted using the subsequent equation [18].

$$\mathrm{PL}{\mathrm{C}}_{\mathrm{i}}=100/(1+\exp \left(\mathrm{a}\right(\mathsf{\Psi }-\mathrm{b}\left)\right))$$

(1)

where PLC_i is the percentage loss of hydraulic conductivity of the stem at each pressurization, Ψ is the xylem water potential, and parameters a and b represent the maximum slope of the curve at 50% loss of hydraulic conductivity and xylem water potential (P₅₀), respectively.

At the completion of the vulnerability curve measurements, the branches were rinsed using a methyI blue (Shanghai McLean Biochemical Technology Co. Ltd., Shanghai, China) solution. Subsequently, the sapwood area (A_S, m²) of each branch was measured to calculate the sapwood-specific conductance (K_S, kg·m⁻¹·s⁻¹·MPa⁻¹). Simultaneously, all leaves on the branch used for measurements were collected individually, these leaves were then scanned using Win RHIZO (Regent Instruments Inc., Quebec City, QC, Canada) to determine the total leaf area (AL, m²) and calculate the leaf-specific conductance (K_L, kg·m⁻¹·s⁻¹·MPa⁻¹). The following formula was used for the calculation of the indicators:

$${\mathrm{K}}_{\mathrm{S}}={\mathrm{K}}_{\mathrm{h}}/{\mathrm{A}}_{\mathrm{S}}$$

(2)

$${\mathrm{K}}_{\mathrm{L}}={\mathrm{K}}_{\mathrm{h}}/{\mathrm{A}}_{\mathrm{L}}$$

(3)

2.5. Statistical Analysis

Differences in plant traits under different water treatments were analyzed using one-way ANOVA in SPSS 26.0 (IBM Corp., Armonk, NY, USA). To examine direct or indirect relationships between C. passerinum traits, we conducted segmented structural equation modeling using the piecewise SEM package in R-4.2.1 (R Foundation for Statistical Computing, Vienna, Austria) [19]. The overall fit of the models was tested using the Fischer’s C statistic [20] and models with low AIC scores were selected. Furthermore, we calculated normalized path coefficients and R² for each path in the model, and each path was treated as independent.

3. Results

3.1. Response of Leaf Water Potential

After the drought treatments, leaf water potential (Ψ_PD and Ψ_MD) was significantly lower in the two drought groups compared with the control, and the values were significantly smaller in the severe drought group than in the mild drought (Figure 1a,b). After rehydration, leaf water potential had a differential change consistent with the above (Figure 1a,b), indicating no significant recovery of leaf water potential in the two drought groups.

3.2. Response of Photosynthetic Characteristics

After drought treatment, Pn was significantly lower in the two drought groups compared with the control, and there was no significant difference between Pn in the mild and severe drought groups. After rehydration, the values of the mild drought group did not change significantly compared with the control, and the values of the severe drought group were significantly lower than the control (Figure 2a), indicating that there was a recovery of Pn in the mild drought group, but not in the severe drought group. Additionally, Tr was significantly lower in the severe drought group compared with the control. After rehydration, there was no significant difference between Tr and control in either of the two drought groups (Figure 2b). Gs was significantly lower in two drought groups compared with the control, but there was no difference between mild and severe. After rehydration, there was no significant difference between the Gs of the two drought groups and the control (Figure 2c), indicating Gs recovery in two drought groups. There was no significant change in WUE in the mild drought group, and a significant increase in WUE in the severe drought group compared with the control. After rehydration, the values were significantly lower in the mild drought group and not significantly different in the severe drought group compared with the control (Figure 2d).

3.3. Response of Hydraulic Conductivity

After drought treatment, there was no significant change in hydraulic conductivity (K_S and K_L) in the two drought groups compared with the control. However, the hydraulic conductivity of the drought group was higher than control, and the mild drought group was higher than the severe drought group. After rehydration, although there was also no significant difference in hydraulic conductivity between the two drought groups and the control, the values for the severe drought group were much lower than the control (Figure 3a,b).

3.4. Response of Embolic Vulnerability

Figure 4a shows the relationship between percentage loss of xylem hydraulic conductivity and water potential, and it was found that the curve for seedlings shifted to the right with increasing drought stress, indicating that xylem embolization increased with increasing drought stress. After rehydration, the curve shifted in accordance with the above observation. After drought treatment, although there was no significant change in P₅₀ in the two drought groups compared with the control, the values increased with increasing drought intensity, indicating weak embolism resistance in stems with stronger drought stress (Figure 4b). After rehydration, the changes in P₅₀ were consistent with those after the drought treatments.

3.5. Relationships among Traits

According to our hypothesized relationships, S. passerine P₅₀ was negatively correlated with K_S, and Gs was positively correlated with Tr (Figure 5). However, the relationships between the other explanatory variables were not supported by the model.

4. Discussion

4.1. Leaf Water Potential and Photosynthetic Characteristics after Drought and Rehydration

In this study, we found that stomatal conductance, transpiration rate, and leaf water potential decreased with increasing drought stress intensity, suggesting that the C. passerinum passes through a series of physiological responses to increase water uptake, and to reduce water dissipation. This result is consistent with the results of most previous studies [21,22,23]. For instance, Toscano et al. found that stomatal conductance of Eugenia uniflora L. ‘Etna Fire’ and P. × fraseri Dress ‘Red Robin’ decreased with increasing drought stress. When plants are subjected to drought stress, a decrease in leaf water potential causes a water deficit in the plant. To prevent excessive water loss, plants lower stomatal conductance and transpiration rate to increase water uptake and thus reduce water loss. However, the reduction in stomatal conductance and transpiration rate limits CO₂ uptake, resulting in an inadequate supply of carbon for photosynthesis. Additionally, water stress negatively affects the functionality of chloroplasts and the enzymatic processes crucial for photosynthesis. As a consequence, the photosynthetic rate decreases as a mechanism to enhance the plant’s adaptability and resistance to drought-stressed environments [24].

When assessing the adaptive capacity of plants to drought, it is crucial to consider not only their performance during the drought period but also their recovery ability after rehydration [25]. This comprehensive evaluation provides a more accurate understanding of a plant’s resilience to drought stress. Absolutely, the ability of plants to quickly recover and compensate for the damage caused by drought stress during rehydration after stress relief serves as a vital indicator of their adaptive capacity to adverse conditions. Here, after rehydration, there was no significant recovery of leaf water potential in the two drought groups. The decrease in plant leaf water potential is caused by embolism [26]. If embolism is not repaired, i.e., there is no significant increase in hydraulic conductivity, the leaf water potential of C. passerinum will not increase, which is to take up more soil water to quickly recover all its physiological properties [27]. However, Gs recovered significantly in two drought groups. This result is consistent with the study by Skelton et al., which found that after experiencing drought-rainfall four shrubs (Leptecophylla divaricata (Hook.f.) C.M.Weiller (Ericaceae), Micrantheum hexandrum Hook.f. (Picrodendraceae), Bursaria spinosa Cav. (Pittosporaceae), and Melaleuca pustulata Hook.f. (Myrtaceae)) were able to recover quickly from gas exchange, suggesting that the shrubs have a strong resistance to embolism, as recovery of gas exchange was predicted by the plants’ high resistance to embolism. Additionally, we found that there was recovery of Pn in the mild drought group, but not in the severe drought group. Reduced activity of enzymes related to plant photosynthesis, damage to the photosynthetic system, and a substantial decrease in the content of photosynthetic pigments were associated with heavy drought stress, and metabolic limitations after rehydration made recovery slower. If the rehydration time is lengthened, the photosynthetic capacity of C. passerinum under heavy drought stress may be fully restored, also suggesting that the plant possesses a hydrodynamic recovery strategy [28]. Notably, there was a temporal difference between the two measurements, and the small sample size for the hydraulic metrics (n = 3) and inconsistent sample size for the photosynthetic indicators may explain the large differences in plant traits between the drought and subsequent rehydration. Additionally, the short rewatering time may have contributed to the lack of significant recovery in C. passerinum traits.

4.2. The Trade-Off between Hydraulic Effectiveness and Safety

After drought treatment, although there were no significant changes in sapwood- and leaf-specific conductance in the two drought groups compared with the control, the values in the drought group were greater than those in the control group, and the mild drought group was greater than the severe drought group, suggesting that drought stress can increase the water use efficiency and leaf water supply capacity of C. passerinum, and that this effect was more pronounced under mild drought stress. This result is consistent with the findings of Zhou et al. [29], who found that sapwood-specific conductance was significantly higher in Haloxylon ammodendron (C. A. Mey.) Bunge under moderate drought stress than in low and high drought stress samples. Compensatory mechanisms may occur in C. passerinum [30]. Mild drought stress causes changes in stem conduit structure, and xylem hydraulic conductivity is affected by the characteristics of the conduit, which improves water use efficiency and water availability [31]. Severe drought stress causes the formation of embolisms in xylem conduits and increased resistance to water use, making water use efficiency lower than in mild drought. After rehydration, although there was also no significant difference between the hydraulic conductivity of the 2 drought groups and the control, the value of the severe drought group was much lower than control, indicating that individuals in the severe drought group were damaged and did not recover. This is because prolonged drought impaired the metabolism of C. passerinum, especially the activity of ribulose bisphosphate carboxylase, preventing rapid recovery of plant hydraulic traits after rewatering [32].

The magnitude of P₅₀ can reflect the embolic resistance of plants [9,33]. Generally, species with low P₅₀ are more resistant to damage from exogenous stresses and plants survive longer [34,35]. We found that after drought treatment, compared with the control, although there was no significant change in embolism resistance in the 2 drought groups, the values increased with the intensity of drought stress, indicating that the stronger the drought stress, the weaker the embolism resistance of the stems. This result is similar to the study by Duan et al. [36]. As drought stress intensifies, the occurrence of cavitation embolism disrupts the integrity of the water column within the xylem conduit. The probability of embolism increases in tandem with the severity of drought, resulting in a concomitant reduction in the plant’s long-distance water transport efficiency and, consequently, compromising its hydraulic resilience and drought tolerance [37]. However, some studies have reported that populations from drier environments are less susceptible to drought than those from wetter environments [38,39]. This discrepancy can be attributed to the fact that the species in this study originate from the same source and exhibit consistent physiological responses to their environment. On the other hand, species that have been distributed in various moisture environments for an extended period have likely adapted to their local conditions, resulting in physiological differences. These variations are examples of intraspecific variation caused by the disparities in environmental conditions. After rehydration, we found that plants in 2 drought groups did not recover. On one hand, it may be because the rehydration time was short and plants could not generate new xylem [12]. On the other hand, non-structural carbon can repair embolized conduits, but long-term drought may have caused carbon starvation [10].

5. Conclusions

The results of the study indicated that increased drought stress had a significant impact on C. passerinum plants, resulting in reduced leaf water potential, stomatal conductance, and impeded net photosynthetic rate. These changes increased the likelihood of xylem embolism occurrence in the plants. We found that drought stress did not significantly enhance water transport in C. passerinum. Meanwhile, it concurrently reduced embolism resistance in the plants. After the drought stress was lifted, the hydraulic conductivity of the severe drought group did not recover to the control level, as did the embolism resistance of the two drought groups, suggesting that a short period of rehydration after drought stress does not allow plants to recovery water transport and embolism resistance. Additionally, significant recovery of stomatal conductance was observed in two drought groups, as well as recovery of Pn in the mild drought group, whereas there was no significant recovery of leaf water potential in two drought groups, because low water potential promotes water uptake by the root system, which in turn promotes the recovery of hydraulic traits.